@article{SU2026, 
author = {Yao SU and Xiuli ZHANG and Hanquan LI and Tao YI and Le XIE and Feng QIU and Yun XIANG and Ziyang ZHOU and Dahua WU},
title = {Molecular mechanism of Zishen Huoxue Formula in improving vascular dementia through GRP78/IRE1/XBP1 signaling pathway mediated endoplasmic reticulum stress},
year = {2026},
journal = {Journal of Beijing University of Traditional Chinese Medicine},
volume = {49},
number = {3},
pages = {341-351},
keywords = {vascular dementia, Zishen Huoxue Formula, hippocampus, endoplasmic reticulum stress, pyroptosis, rats},
url = {https://www.sciopen.com/article/10.3969/j.issn.1006-2157.2026.03.006},
doi = {10.3969/j.issn.1006-2157.2026.03.006},
abstract = {ObjectiveTo investigate the mechanism by which Zishen Huoxue Formula (ZSHXF) regulates the glucose-regulated protein 78 (GRP78)/inositol-requiring enzyme 1 (IRE1)/X-box Binding Protein 1 (XBP1) signaling pathway to inhibit hippocampal neuronal pyroptosis and improve cognitive function in vascular dementia (VD) rats.MethodsA total of seventy-two Sprague-Dawley (SD) rats were randomly allocated into six groups (n=12 per group) based on random number table: a Sham group, a Model group, a Donepezil hydrochloride group (0.45 mg/kg), and three ZSHXF groups designated as low-dose (8.9 g/kg), medium-dose (17.8 g/kg), and high-dose (35.6 g/kg). VD models were induced in all groups, with the exception of the sham group, which underwent blunt dissection without ligation of the bilateral common carotid arteries. Following the modeling procedure, the Donepezil hydrochloride and ZSHXF groups received daily oral gavage for 14 consecutive days. In contrast, the sham and model groups were administered an equivalent volume of normal saline via oral gavage.The spatial learning, memory, and exploratory capacities of rats were evaluated through the application of the Morris Water Maze (MWM) test and the Y-maze test. HE staining was performed to assess morphological alterations in rat hippocampal CA1 region neurons. The ultrastructural characteristics of the endoplasmic reticulum within hippocampal neurons were investigated using Transmission Electron Microscopy (TEM). Immunofluorescence staining was conducted to evaluate fluorescence co-localization intensity of GRP78 with the neuronal nuclei antigen (NeuN), and gasdermin D (GSDMD) with βⅢ Tubulin in hippocampal neurons. The expression levels of GRP78, phosphorylated IRE1 (p-IRE1), IRE1, XBP1, CCAAT/enhancer-binding protein homologous protein (CHOP), NOD-like receptor protein 3 (NLRP3), GSDMD and cysteine-aspartic acid protease 1 (Caspase-1) in the rat hippocampus were quantified using Western blot analysis.ResultsIn comparison to the sham group, the model group demonstrated a prolonged escape latency (P < 0.01), a reduction in both platform crossings and the duration spent in the target quadrant (P < 0.01), and a decreased spontaneous alternation rate in the Y-maze (P < 0.01). Additionally, there was an increase in hippocampal CA1 region neuronal damage, evident dilation and swelling of the endoplasmic reticulum, and enhanced fluorescence co-localization expression of GRP78 with NeuN and GSDMD with βⅢ Tubulin in the rat hippocampus. Furthermore, elevated expression levels of GRP78, p-IRE1/IRE1, XBP1, CHOP, NLRP3, GSDMD, and Caspase-1 were observed in hippocampus (P < 0.01). Compared to the model group, the donepezil hydrochloride group and low- and medium- and high-dose ZSHXF groups had shorter escape latency (P < 0.01), an increase in platform crossings and time spent in the target quadrant (P < 0.05, P < 0.01), and an improvement in the spontaneous alternation rate in the Y-maze (P < 0.05, P < 0.01). This intervention also alleviated hippocampal CA1 region neuronal damage, inhibited the dilation and swelling of the endoplasmic reticulum lumen, restored the number of endoplasmic reticulum, reduced the fluorescence co-localization intensity of GRP78 with NeuN and GSDMD with βⅢ Tubulin in the hippocampus; The donepezil hydrochloride group showed reduced GSDMD expression (P < 0.01). All dose groups of ZSHXF exhibited decreased protein expression levels of XBP1, CHOP, NLRP3, GSDMD, and Caspase-1 (P < 0.05, P < 0.01). Additionally, the medium- and high-dose groups of ZSHXF reduced GRP78 protein expression and a decreased p-IRE1/IRE1 ratio (P < 0.05, P < 0.01). ConclusionZSHXF markedly mitigates hippocampal damage and enhances learning and memory capabilities in rats model subjected to VD. This neuroprotective effect is potentially linked to the modulation of the endoplasmic reticulum stress, specifically through the GRP78/IRE1/XBP1 signaling pathway, which subsequently inhibits hippocampal neuronal pyroptosis.}
}